The overall goal of the research objectives in this proposal is to understand the role of hyaluronan (HA) and HA-binding proteins in the proliferative and morphogenic processes that occur in normal blood vessel development as well as during vessel wall injury repair. We previously cloned and characterized a serum-inducible, secreted HA- binding protein in SMCs called TSG-6. The expression of TSG-6 in cultured SMCs is tightly controlled by growth factors and cytokines and, in vivo, TSG-6 expression is specifically associated with proliferating, neointimal SMCs in the injured blood vessel and with activated macrophages, modified SMCs, and microvessel endothelial cells in human atherectomy samples. We have also observed a number of striking effects in SMCs constitutively overexpressing TSG-6: First, an increase in cell size was observed in all the independent clonal lines we examined and in half of these cell lines the hypertrophic response size was observed in all the independent clonal lines we examined and in half of these cell lines the hypertrophic response was accompanied by a shift in DNA content from diploid to tetraploid. Second, cell lines that expressed high levels of TSG-6 also exhibited an increased growth rate. Finally, these cells were able to elaborate large bundles of mature collagen fibrils in culture. In this application, we will examine the process by which TSG-6 mediates SMC hypertrophy, hyperplasia and, collagen fibrillogenesis. We propose to test the hypothesis that the observed growth-related changes in TSG-6 transfected cells are due to the ability of this protein to influence collagen fibrillogenesis. We will determine whether our observation is related to the fact that the carboxy1-terminal proximal domain of TSG-6 exhibits sequence similarity with a protein that binds to procollagen C-propeptide and enhances collagen fibrillogenesis. It is also known that vasoconstrictors such as angiotensin II, thromboxane A2, endothelin and norepinephrine are able to act directly on SMCs and induce a hypertrophic response. We will test the hypothesis that TSG-6 maybe a downstream effector molecule for these hypertensive agents. The specific aims are: 1. To characterize the cellular responses (e.g. hypertrophy, hyperplasia, and enhanced collagen fibrillogenesis) mediated by the HA-binding protein TSG-6. 2. To identify the molecular mechanism(s) responsible for TSG-6 mediated increase in collagen fibrillogenesis. 3. To determine whether TSG-6 is a downstream effector molecule of hypertrophic agents such as angiotensin II and its putative mediator, TGF-beta1. The proposed studies will provide a better understanding of the role of TSG-6 in the vascular healing process.